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Abstract:

The present invention relates to therapeutic combinations of
[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid
diisopropoxycarbonyloxymethyl ester (tenofovir disoproxil fumarate,
Viread®) and
(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one (emtricitabine, Emtriva®, (-)-cis FTC) and their
physiologically functional derivatives. The combinations may be useful in
the treatment of HIV infections, including infections with HIV mutants
bearing resistance to nucleoside and/or non-nucleoside inhibitors. The
present invention is also concerned with pharmaceutical compositions and
formulations of said combinations of tenofovir disoproxil fumarate and
emtricitabine, and their physiologically functional derivatives, as well
as therapeutic methods of use of those compositions and formulations.

Claims:

1. A method for the treatment or prevention of the symptoms or effects of
an HIV infection in an infected animal which comprises administering to
said animal a therapeutically effective amount of a composition
comprising [2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid
diisopropoxycarbonyloxymethyl ester fumarate (tenofovir disoproxil
fumarate) or a physiologically functional derivative thereof, and
(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one (emtricitabine) or a physiologically functional
derivative thereof.

Description:

[0001] This non-provisional application is a continuation of U.S. patent
application Ser. No. 12/204,174, filed Sep. 4, 2008, which is a
continuation of U.S. patent application Ser. No. 10/540,794, filed Mar.
20, 2006, which is a national stage entry of PCT/US04/00832, filed Jan.
13, 2004 which claims the benefit of Provisional Application Nos.
60/440,246 and 60/440,308, both filed Jan. 14, 2003, the disclosures of
each of which is incorporated herein by reference.

FIELD OF THE INVENTION

[0002] The invention relates generally to combinations of compounds with
antiviral activity and more specifically with anti-HIV properties. In
particular, it relates to chemically stable combinations of structurally
diverse anti-viral agents.

BACKGROUND OF THE INVENTION

[0003] Human immunodeficiency virus (HIV) infection and related diseases
are a major public health problem worldwide. Human immunodeficiency virus
type 1 (HIV-1) encodes at least three enzymes which are required for
viral replication: reverse transcriptase (RT), protease (Prt), and
integrase (Int). Although drugs targeting reverse transcriptase and
protease are in wide use and have shown effectiveness, particularly when
employed in combination, toxicity and development of resistant strains
have limited their usefulness (Palella, et al N. Engl. J. Med. (1998)
338:853-860; Richman, D. D. Nature (2001) 410:995-1001). Human
immunodeficiency virus type 1 (HIV-1) protease (Prt) is essential for
viral replication and is an effective target for approved antiviral
drugs. The HIV Prt cleaves the viral Gag and Gag-Pol polyproteins to
produce viral structural proteins (p17, p24, p7 and p6) and the three
viral enzymes. Combination therapy with RT inhibitors has proven to be
highly effective in suppressing viral replication to unquantifiable
levels for a sustained period of time. Also, combination therapy with RT
and Prt inhibitors (PI) have shown synergistic effects in suppressing HIV
replication. Unfortunately, a high percentage, typically 30 to 50% of
patients currently fail combination therapy due to the development of
drug resistance, non-compliance with complicated dosing regimens,
pharmacokinetic interactions, toxicity, and lack of potency. Therefore,
there is a need for new HIV-1 inhibitors that are active against mutant
HIV strains, have distinct resistance profiles, fewer side effects, less
complicated dosing schedules, and are orally active. In particular, there
is a need for a less onerous dosage regimen, such as once per day oral
dosing, optimally with as few pills as possible.

[0004] The use of combinations of compounds can yield an equivalent
antiviral effect with reduced toxicity, or an increase in drug efficacy.
Lower overall drug doses can reduce the frequency of occurrence of
drug-resistant variants of HIV. Many different methods have been used to
examine the effects of combinations of compounds acting together in
different assay systems (Furman WO 02/068058). Lower doses predict better
patient compliance when pill burden decreases, dosing schedules are
simplified and, optionally, if synergy between compounds occurs (Loveday,
C. "Nucleoside reverse transcriptase inhibitor resistance" (2001) JAIDS
Journal of Acquired Immune Deficiency Syndromes 26:S10-S24). AZT
(Zidovudine®, 3'-azido, 3'-deoxythymidine) demonstrates synergistic
antiviral activity in vitro in combination with agents that act at HIV-1
replicative steps other than reverse transcription, including recombinant
soluble CD4 castanospermine and recombinant interferon-α. However,
it must be noted that combinations of compounds can give rise to
increased cytotoxicity. For example, AZT and recombinant
interferon-α have an increased cytotoxic effect on normal human
bone marrow progenitor cells.

[0005] Chemical stability of combinations of antiviral agents is an
important aspect of co-formulation success and the present invention
provides examples of such combinations.

[0006] There is a need for new combinations of orally-active drugs for the
treatment of patients infected with certain viruses, e.g. HIV, that
provide enhanced therapeutic safety and efficacy, impart lower
resistance, and predict higher patient compliance.

SUMMARY OF THE INVENTION

[0007] The present invention provides combinations of antiviral compounds,
in particular compositions and methods for inhibition of HIV. In an
exemplary aspect, the invention includes a composition including
tenofovir disoproxil fumarate and emtricitabine which has anti-HIV
activity. The composition of tenofovir DF and emtricitabine is both
chemically stable and either synergistic and/or reduces the side effects
of one or both of tenofovir DF and emtricitabine. Increased patient
compliance is likely in view of the lower pill burden and simplified
dosing schedule.

[0008] The present invention relates to therapeutic combinations of
[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid
diisopropoxycarbonyloxymethyl ester fumarate (tenofovir disoproxil
fumarate, tenofovir DF, TDF, Viread®) and
(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one (emtricitabine, Emtriva®, (-)-cis FTC) and their use
in the treatment of HIV infections including infections with HIV mutants
bearing resistance to nucleoside and/or non-nucleoside inhibitors. The
present invention is also concerned with pharmaceutical compositions and
formulations of said combinations of tenofovir disoproxil fumarate and
emtricitabine. Another aspect of the invention is a pharmaceutical
formulation comprising a physiologically functional derivative of
tenofovir disoproxil fumarate or a physiologically functional derivative
of emtricitabine.

[0009] Therapeutic combinations and pharmaceutical compositions and
formulations of the invention include the combination of PMEA or PMPA
(tenofovir) compounds with emtricitabine or
(2R,5S,cis)-4-amino-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-pyrimidi-
n-2-one (3TC, lamivudine, Epivir®), and their use in the treatment of
HIV infections.

[0010] One aspect of the invention is a method for the treatment or
prevention of the symptoms or effects of an HIV infection in an infected
animal which comprises administering to, i.e. treating, said animal with
a therapeutically effective amount of a combination comprising
[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid
diisopropoxycarbonyloxymethyl ester fumarate (tenofovir DF, TDF) or a
physiologically functional derivative thereof, and
(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one (emtricitabine) or a physiologically functional
derivative thereof.

[0011] Another aspect of the invention is a unit dosage form of a
therapeutic combination comprising tenofovir disoproxil fumarate and
emtricitabine, or physiological functional derivatives thereof. The unit
dosage form may be formulated for administration by oral or other routes
and is unexpectedly chemically stable in view of the properties of the
structurally diverse components.

[0012] Another aspect of the invention is directed to chemically stable
combination antiviral compositions comprising tenofovir disoproxil
fumarate and emtricitabine. In a further aspect of the invention, the
chemically stable combinations of tenofovir disoproxil fumarate and
emtricitabine further comprise a third antiviral agent. In this
three-component mixture, the unique chemical stability of tenofovir
disoproxil fumarate and emtricitabine is taken advantage of in order to
enable the combination with the third antiviral agent. Particularly
useful third agents include, by way of example and not limitation, those
of Table A. Preferably, the third component is an agent approved for
antiviral use in humans, more preferably, it is an NNRTI or a protease
inhibitor (PI), more preferably yet, it is an NNRTI. In a particularly
preferred embodiment, the invention is directed to a combination of the
chemically stable mixture of tenofovir disoproxil fumarate and
emtricitabine together with efavirenz.

[0013] Another aspect of the invention is a patient pack comprising at
least one, typically two, and optionally, three active ingredients and
other antiviral agents selected from tenofovir disoproxil fumarate and
emtricitabine, and an information insert containing directions on the use
of tenofovir disoproxil fumarate and emtricitabine together in
combination.

[0014] Another aspect of the invention is a process for preparing the
combinations hereinbefore described, which comprises bringing into
association tenofovir DF and emtricitabine of the combination in a
medicament to provide an antiviral effect. In a further aspect of the
present invention, there is provided the use of a combination of the
present invention in the manufacture of a medicament for the treatment of
any of the aforementioned viral infections or conditions.

DETAILED DESCRIPTION OF THE INVENTION

[0015] While the invention will be described in conjunction with the
enumerated claims, it will be understood that they are not intended to
limit the invention to those claims. On the contrary, the invention is
intended to cover all alternatives, modifications, and equivalents, which
may be included within the scope of the present invention as defined by
the claims.

DEFINITIONS

[0016] Unless stated otherwise, the following terms and phrases as used
herein are intended to have the following meanings:

[0017] When tradenames are used herein, applicants intend to independently
include the tradename product and the active pharmaceutical ingredient(s)
of the tradename product.

[0018] The term "chemical stability" means that the two primary antiviral
agents in combination are substantially stable to chemical degradation.
Preferably, they are sufficiently stable in physical combination to
permit commercially useful shelf life of the combination product.
Typically, "chemically stable" means that a first component of the
mixture does not act to degrade a second component when the two are
brought into physical combination to form a pharmaceutical dosage form.
More typically, "chemically stable" means that the acidity of a first
component does not catalyzes or otherwise accelerate the acid
decomposition of a second component. By way of example and not
limitation, in one aspect of the invention, "chemically stable" means
that tenofovir disoproxil fumarate is not substantially degraded by the
acidity of emtricitabine. "Substantially" in this context means at least
about less than 10%, preferably less than 1%, more preferably less than
0.1%, more preferably yet, less than 0.01% acid degradation of tenofovir
disoproxil fumarate over a 24-hour period when the products are in a
pharmaceutical dosage form.

[0019] The terms "synergy" and "synergistic" mean that the effect achieved
with the compounds used together is greater than the sum of the effects
that results from using the compounds separately, i.e. greater than what
would be predicted based on the two active ingredients administered
separately. A synergistic effect may be attained when the compounds are:
(1) co-formulated and administered or delivered simultaneously in a
combined formulation; (2) delivered by alternation or in parallel as
separate formulations; or (3) by some other regimen. When delivered in
alternation therapy, a synergistic effect may be attained when the
compounds are administered or delivered sequentially, e.g. in separate
tablets, pills or capsules, or by different injections in separate
syringes. In general, during alternation therapy, an effective dosage of
each active ingredient is administered sequentially, i.e. serially,
whereas in combination therapy, effective dosages of two or more active
ingredients are administered together. A synergistic antiviral effect
denotes an antiviral effect which is greater than the predicted purely
additive effects of the individual compounds of the combination.

[0020] The term "physiologically functional derivative" means a
pharmaceutically active compound with equivalent or near equivalent
physiological functionality to tenofovir DF or emtricitabine when
administered in combination with another pharmaceutically active compound
in a combination of the invention. As used herein, the term
"physiologically functional derivative" includes any: physiologically
acceptable salt, ether, ester, prodrug, solvate, stereoisomer including
enantiomer, diastereomer or stereoisomerically enriched or racemic
mixture, and any other compound which upon administration to the
recipient, is capable of providing (directly or indirectly) such a
compound or an antivirally active metabolite or residue thereof.

[0021] "Bioavailability" is the degree to which the pharmaceutically
active agent becomes available to the target tissue after the agent's
introduction into the body. Enhancement of the bioavailability of a
pharmaceutically active agent can provide a more efficient and effective
treatment for patients because, for a given dose, more of the
pharmaceutically active agent will be available at the targeted tissue
sites.

[0022] The compounds of the combinations of the invention may be referred
to as "active ingredients" or "pharmaceutically active agents."

[0023] The term "prodrug" as used herein refers to any compound that when
administered to a biological system generates the drug substance, i.e.
active ingredient, as a result of spontaneous chemical reaction(s),
enzyme catalyzed chemical reaction(s), and/or metabolic chemical
reaction(s).

[0024] "Prodrug moiety" means a labile functional group which separates
from the active inhibitory compound during metabolism, systemically,
inside a cell, by hydrolysis, enzymatic cleavage, or by some other
process (Bundgaard, Hans, "Design and Application of Prodrugs" in
Textbook of Drug Design and Development (1991), P. Krogsgaard-Larsen and
H. Bundgaard, Eds. Harwood Academic Publishers, pp. 113-191). Prodrug
moieties can serve to enhance solubility, absorption and lipophilicity to
optimize drug delivery, bioavailability and efficacy. A "prodrug" is thus
a covalently modified analog of a therapeutically-active compound.

[0026] "Aryl" means a monovalent aromatic hydrocarbon radical of 6-20
carbon atoms derived by the removal of one hydrogen atom from a single
carbon atom of a parent aromatic ring system. Typical aryl groups
include, but are not limited to, radicals derived from benzene,
substituted benzene, naphthalene, anthracene, biphenyl, and the like.

[0027] "Arylalkyl" refers to an acyclic alkyl radical in which one of the
hydrogen atoms bonded to a carbon atom, typically a terminal or sp3
carbon atom, is replaced with an aryl radical. Typical arylalkyl groups
include, but are not limited to, benzyl, 2-phenylethan-1-yl,
2-phenylethen-1-yl, naphthylmethyl, 2-naphthylethan-1-yl,
2-naphthylethen-1-yl, naphthobenzyl, 2-naphthophenylethan-1-yl and the
like. The arylalkyl group 6 to 20 carbon atoms e.g., the alkyl moiety,
including alkanyl, alkenyl or alkynyl groups, of the arylalkyl group is 1
to 6 carbon atoms and the aryl moiety is 5 to 14 carbon atoms.

[0030] Stereochemical definitions and conventions used herein generally
follow S. P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984)
McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S.,
Stereochemistry of Organic Compounds (1994) John Wiley & Sons, Inc., New
York. Many organic compounds exist in optically active forms, i.e., they
have the ability to rotate the plane of plane-polarized light. In
describing an optically active compound, the prefixes D and L or R and S
are used to denote the absolute configuration of the molecule about its
chiral center(s). The prefixes d and l or (+) and (-) are employed to
designate the sign of rotation of plane-polarized light by the compound,
with (-) or l meaning that the compound is levorotatory. A compound
prefixed with (+) or d is dextrorotatory. For a given chemical structure,
these compounds, called stereoisomers, are identical except that they are
mirror images of one another. A specific stereoisomer is also referred to
as an enantiomer, and a mixture of such isomers is often called an
enantiomeric mixture. A 50:50 mixture of enantiomers is referred to as a
racemic mixture or a racemate. The terms "racemic mixture" and "racemate"
refer to an equimolar mixture of two enantiomeric species, devoid of
optical activity.

[0031] The term "chiral" refers to molecules which have the property of
non-superimposability of the mirror image partner, while the term
"achiral" refers to molecules which are superimposable on their mirror
image partner.

[0032] The term "stereoisomers" refers to compounds which have identical
chemical constitution, but differ with regard to the arrangement of the
atoms or groups in space.

[0033] "Diastereomer" refers to a stereoisomer with two or more centers of
chirality and whose molecules are not mirror images of one another.
Diastereomers have different physical properties, e.g. melting points,
boiling points, spectral properties, and reactivities. Mixtures of
diastereomers may separate under high resolution analytical procedures
such as electrophoresis and chromatography.

[0034] "Enantiomers" refer to two stereoisomers of a compound which are
non-superimposable mirror images of one another.

Active Ingredients of the Combinations

[0035] The present invention provides novel combinations of two or more
active ingredients being employed together. In some embodiments, a
synergistic antiviral effect is achieved. In other embodiments, a
chemically stable combination is obtained. The combinations include at
least one active ingredient selected from (1) tenofovir disoproxil
fumarate and physiologically functional derivatives, and at least one
active ingredient selected from (2) emtricitabine and physiologically
functional derivatives. The term "synergistic antiviral effect" is used
herein to denote an antiviral effect which is greater than the predicted
purely additive effects of the individual components (a) and (b) of the
combination.

[0038] The chemical names for Tenofovir disoproxil include:
[2-(6-amino-purin-9-yl)-1-methyl-ethoxymethyl]-phosphonic acid
diisopropoxycarbonyloxymethyl ester;
9-[(R)-2-[[bis[[(isopropoxycarbonyl)oxy]methoxy]phosphinyl]methoxy]propyl-
]adenine; and 2,4,6,8-tetraoxa-5-phosphanonanedioic acid,
5-[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]-,
bis(1-methylethyl)ester, 5-oxide. The CAS Registry numbers include:
201341-05-1; 202138-50-9; 206184-49-8. It should be noted that the
ethoxymethyl unit of tenofovir has a chiral center. The R (rectus, right
handed configuration) enantiomer is shown. However, the invention also
includes the S isomer. The invention includes all enantiomers,
diastereomers, racemates, and enriched stereoisomer mixtures of tenofovir
(PMPA) and physiologically functional derivatives thereof.

[0040] The chemical names of PMPA, tenofovir include:
(R)-9-(2-phosphonylmethoxypropyl)adenine; and phosphonic acid,
[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]. The CAS Registry
number is 147127-20-6.

[0041] Tenofovir disoproxil fumarate (DF) is a nucleotide reverse
transcriptase inhibitor approved in the United States in 2001 for the
treatment of HIV-1 infection in combination with other antiretroviral
agents. Tenofovir disoproxil fumarate or Viread® (Gilead Science,
Inc.) is the fumarate salt of tenofovir disoproxil. Viread® may be
named as: 9-[(R)-2-[[bis[[(isopropoxycarbonyl)oxy]methoxy]phosphinyl]meth-
oxy]propyl]adenine fumarate (1:1); or
2,4,6,8-tetraoxa-5-phosphanonanedioic acid,
5-[[(1R)-2-(6-amino-9H-purin-9-yl)-1-methylethoxy]methyl]-,
bis(1-methylethyl)ester, 5-oxide, (2E)-2-butenedioate (1:1). The CAS
Registry number is 202138-50-9.

[0042] Physiologically functional derivatives of tenofovir disoproxil
fumarate include PMEA (adefovir,
9-((R)-2-(phosphonomethoxy)ethyl)adenine) and PMPA compounds. Exemplary
combinations include a PMEA or PMPA compound in combination with
emtricitabine or 3TC. PMEA and PMPA compounds have the structures:

[0043] The PMPA compound may be enantiomerically-enriched or purified
(single stereoisomer) where the carbon atom bearing R3 may be the R
or S enantiomer. The PMPA compound may be a racemate, i.e. a mixture of R
and S stereoisomers.

[0047] The chemical names for emtricitabine include: (-)-cis-FTC;
β'-L-hydroxymethyl-5-(5-fluorocytosin-1-yl)-1,3-oxathiolane;
(2R,5S)-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine; and
4-amino-5-fluoro-1-(2-hydroxymethyl-[1,3]-(2R,5S)-oxathiolan-5-yl)-1H-pyr-
imidin-2-one. The CAS Registry numbers include: 143491-57-0; 143491-54-7.
It should be noted that FTC contains two chiral centers, at the 2 and 5
positions of the oxathiolane ring, and therefore can exist in the form of
two pairs of optical isomers (i.e. enantiomers) and mixtures thereof
including racemic mixtures. Thus, FTC may be either a cis or a trans
isomer or mixtures thereof. Mixtures of cis and trans isomers are
diastereomers with different physical properties. Each cis and trans
isomer can exist as one of two enantiomers or mixtures thereof including
racemic mixtures. The invention includes all enantiomers, diastereomers,
racemates, and enriched stereoisomer mixtures of emtricitabine and
physiologically functional derivatives thereof. For example, the
invention includes physiological functional derivatives such as the 1:1
racemic mixture of the enantiomers
(2R,5S,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one (emtricitabine) and its mirror image
(2S,5R,cis)-4-amino-5-fluoro-1-(2-hydroxymethyl-1,3-oxathiolan-5-yl)-(1H)-
-pyrimidin-2-one, or mixtures of the two enantiomers in any relative
amount. The invention also includes mixtures of cis and trans forms of
FTC.

[0048] Physiologically functional derivatives of emtricitabine include 1,3
oxathiolane nucleosides having the structure:

[0050] Nucleobases B may be attached in the configurations of
naturally-occurring nucleic acids to the 1,3 oxathiolane moiety through a
covalent bond between the N-9 of purines, e.g. adenin-9-yl and
guanin-9-yl, or N-1 of pyrimidines, e.g. thymin-1-yl and cytosin-1-yl
(Blackburn, G. and Gait, M. Eds. "DNA and RNA structure" in Nucleic Acids
in Chemistry and Biology, 2nd Edition, (1996) Oxford University
Press, pp. 15-81).

[0052] Physiologically functional derivatives of emtricitabine also
include 3TC (lamivudine, Epivir®), a reverse transcriptase inhibitor
approved in the United States for the treatment of HIV-1 infection in
combination with AZT as Combivir® (GlaxoSmithKline). U.S. Pat. Nos.
5,859,021; 5,905,082; 6,177,435; 5,627,186; 6,417,191. Lamivudine (U.S.
Pat. Nos. 5,587,480, 5,696,254, 5,618,820, 5,756,706, 5,744,596,
5,681,64, 5,466,806, 5,151,426) has the structure:

##STR00006##

[0053] For example and for some therapeutic uses, 3TC may be a
physiologically functional derivative of emtricitabine in combination
with tenofovir DF or a physiologically functional derivative of tenofovir
DF.

[0054] It will be appreciated that tenofovir DF and emtricitabine, and
their physiologically functional derivatives may exist in keto or enol
tautomeric forms and the use of any tautomeric form thereof is within the
scope of this invention. Tenofovir DF and emtricitabine will normally be
utilized in the combinations of the invention substantially free of the
corresponding enantiomer, that is to say no more than about 5% w/w of the
corresponding enantiomer will be present.

Prodrugs

[0055] The invention includes all prodrugs of tenofovir and emtricitabine.
An exemplary prodrug of tenofovir is tenofovir disoproxil fumarate (TDF,
Viread@). A large number of structurally-diverse prodrugs have been
described for phosphonic acids (Freeman and Ross in Progress in Medicinal
Chemistry 34: 112-147 (1997). A commonly used prodrug class is the
acyloxyalkyl ester, which was first used as a prodrug strategy for
carboxylic acids and then applied to phosphates and phosphonates by
Farquhar et al (1983) J. Pharm. Sci. 72: 324; also U.S. Pat. Nos.
4,816,570, 4,968,788, 5,663,159 and 5,792,756. Subsequently, the
acyloxyalkyl ester was used to deliver phosphonic acids across cell
membranes and to enhance oral bioavailability. A close variant of the
acyloxyalkyl ester strategy, the alkoxycarbonyloxyalkyl ester, may also
enhance oral bioavailability as a prodrug moiety in the compounds of the
combinations of the invention. Aryl esters of phosphorus groups,
especially phenyl esters, are reported to enhance oral bioavailability
(DeLambert et al (1994) J. Med. Chem. 37: 498). Phenyl esters containing
a carboxylic ester ortho to the phosphate have also been described
(Khamnei and Torrence, (1996) J. Med. Chem. 39:4109-4115). Benzyl esters
are reported to generate the parent phosphonic acid. In some cases,
substituents at the ortho- or para-position may accelerate the
hydrolysis. Benzyl analogs with an acylated phenol or an alkylated phenol
may generate the phenolic compound through the action of enzymes, e.g.
esterases, oxidases, etc., which in turn undergoes cleavage at the
benzylic C--O bond to generate the phosphoric acid and the quinone
methide intermediate. Examples of this class of prodrugs are described by
Mitchell et al (1992) J. Chem. Soc. Perkin Trans. I 2345; Brook et al WO
91/19721. Still other benzylic prodrugs have been described containing a
carboxylic ester-containing group attached to the benzylic methylene
(Glazier et al WO 91/19721). Thio-containing prodrugs are reported to be
useful for the intracellular delivery of phosphonate drugs. These
proesters contain an ethylthio group in which the thiol group is either
esterified with an acyl group or combined with another thiol group to
form a disulfide. Deesterification or reduction of the disulfide
generates the free thio intermediate which subsequently breaks down to
the phosphoric acid and episulfide (Puech et al (1993) Antiviral Res.,
22: 155-174; Benzaria et al (1996) J. Med. Chem. 39: 4958). Cyclic
phosphonate esters have also been described as prodrugs of
phosphorus-containing compounds.

[0056] Prodrug esters in accordance with the invention are independently
selected from the following groups: (1) mono-, di-, and tri-phosphate
esters of tenofovir or emtricitabine or any other compound which upon
administration to a human subject is capable of providing (directly or
indirectly) said mono-, di, or triphosphate ester; (2) carboxylic acid
esters (3) sulphonate esters, such as alkyl- or aralkylsulphonyl (for
example, methanesulphonyl); (4) amino acid esters (for example, alanine,
L-valyl or L-isoleucyl); (5) phosphonate; and (6) phosphonamidate esters.

[0059] Pharmaceutically acceptable prodrugs refer to a compound that is
metabolized in the host, for example hydrolyzed or oxidized, by either
enzymatic action or by general acid or base solvolysis, to form an active
ingredient. Typical examples of prodrugs of the active ingredients of the
combinations of the invention have biologically labile protecting groups
on a functional moiety of the active compound. Prodrugs include compounds
that can be oxidized, reduced, aminated, deaminated, esterified,
deesterified, alkylated, dealkylated, acylated, deacylated,
phosphorylated, dephosphorylated, or other functional group change or
conversion involving forming or breaking chemical bonds on the prodrug.

Chemical Stability of a Pharmaceutical Formulation

[0060] The chemical stability of the active ingredients in a
pharmaceutical formulation is of concern to minimize the generation of
impurities and ensure adequate shelf-life. The active ingredients,
tenofovir disoproxil fumarate and emtricitabine, in the pharmaceutical
formulations of the invention have relatively low pKa values, indicative
of the potential to cause acidic hydrolysis of the active ingredients.
Emtricitabine, with a pKa of 2.65 (Emtriva® Product Insert, Gilead
Sciences, Inc. 2003, available at gilead.com) is subject to hydrolytic
deamination of the 5-fluoro cytosine nucleobase to form the 5-fluoro
uridine nucleobase. Tenofovir disoproxil fumarate, with a pKa of 3.75
(Yuan L. et al "Degradation Kinetics of Oxycarbonyloxymethyl Prodrugs of
Phosphonates in Solution", Pharmaceutical Research (2001) Vol. 18, No. 2,
234-237), is subject also to hydrolytic deamination of the exocyclic
amine of the adenine nucleobase, and to hydrolysis of one or both of the
POC ester groups (U.S. Pat. No. 5,922,695). It is desirable to formulate
a therapeutic combination of tenofovir disoproxil fumarate and
emtricitabine, and the physiological functional derivatives thereof, with
a minimum of impurities and adequate stability.

[0061] The combinations of the present invention provide combination
pharmaceutical dosage forms which are chemically stable to acid
degradation of: (1) a first component (such as tenofovir disoproxil
fumarate, and physiological functional derivatives; (2) a second
component (such as emtricitabine, and physiological functional
derivatives; and (3) optionally a third component having antiviral
activity. The third component includes anti-HIV agents and include:
protease inhibitors (PI), nucleoside reverse transcriptase inhibitors
(NRTI), non-nucleoside reverse transcriptase inhibitors (NNRTI), and
integrase inhibitors. Exemplary third active ingredients to be
administered in combination with first and second components are shown in
Table A. First and second components are as defined in the above section
entitled: ACTIVE INGREDIENTS OF THE COMBINATIONS.

Salts

[0062] Any reference to any of the compounds in the compositions of the
invention also includes any physiologically acceptable salt thereof.
Examples of physiologically acceptable salts of tenofovir DF,
emtricitabine and their physiologically functional derivatives include
salts derived from an appropriate base, such as an alkali metal (for
example, sodium), an alkaline earth (for example, magnesium), ammonium
and NX4.sup.+ (wherein X is C1-C4 alkyl), or an organic
acid such as fumaric acid, acetic acid, succinic acid. Physiologically
acceptable salts of an hydrogen atom or an amino group include salts of
organic carboxylic acids such as acetic, benzoic, lactic, fumaric,
tartaric, maleic, malonic, malic, isethionic, lactobionic and succinic
acids; organic sulfonic acids, such as methanesulfonic, ethanesulfonic,
benzenesulfonic and p-toluenesulfonic acids; and inorganic acids, such as
hydrochloric, sulfuric, phosphoric and sulfamic acids. Physiologically
acceptable salts of a compound of an hydroxy group include the anion of
said compound in combination with a suitable cation such as Na.sup.+ and
NX4.sup.+ (wherein X is independently selected from H or a
C1-C4 alkyl group).

[0063] For therapeutic use, salts of active ingredients of the
combinations of the invention will be physiologically acceptable, i.e.
they will be salts derived from a physiologically acceptable acid or
base. However, salts of acids or bases which are not physiologically
acceptable may also find use, for example, in the preparation or
purification of a physiologically acceptable compound. All salts, whether
or not derived from a physiologically acceptable acid or base, are within
the scope of the present invention.

Administration of the Formulations

[0064] While it is possible for the active ingredients of the combination
to be administered alone and separately as monotherapies, it is
preferable to administer them as a pharmaceutical co-formulation. A
two-part or three-part combination may be administered simultaneously or
sequentially. When administered sequentially, the combination may be
administered in one, two, or three administrations.

[0065] Preferably, two-part or three-part combinations are administered in
a single pharmaceutical dosage form. More preferably, a two-part
combination is administered as a single oral dosage form and a three-part
combination is administered as two identical oral dosage forms. Examples
include a single tablet of tenofovir disoproxil fumarate and
emtricitabine, or two tablets of tenofovir disoproxil fumarate,
emtricitabine, and efavirenz.

[0066] It will be appreciated that the compounds of the combination may be
administered: (1) simultaneously by combination of the compounds in a
co-formulation or (2) by alternation, i.e. delivering the compounds
serially, sequentially, in parallel or simultaneously in separate
pharmaceutical formulations. In alternation therapy, the delay in
administering the second, and optionally a third active ingredient,
should not be such as to lose the benefit of a synergistic therapeutic
effect of the combination of the active ingredients. By either method of
administration (1) or (2), ideally the combination should be administered
to achieve peak plasma concentrations of each of the active ingredients.
A one pill once-per-day regimen by administration of a combination
co-formulation may be feasible for some HIV-positive patients. Effective
peak plasma concentrations of the active ingredients of the combination
will be in the range of approximately 0.001 to 100 μM. Optimal peak
plasma concentrations may be achieved by a formulation and dosing regimen
prescribed for a particular patient. It will also be understood that
tenofovir DF and emtricitabine, or the physiologically functional
derivatives of either thereof, whether presented simultaneously or
sequentially, may be administered individually, in multiples, or in any
combination thereof. In general, during alternation therapy (2), an
effective dosage of each compound is administered serially, where in
co-formulation therapy (1), effective dosages of two or more compounds
are administered together.

Formulation of the Combinations

[0067] When the individual components of the combination are administered
separately they are generally each presented as a pharmaceutical
formulation. The references hereinafter to formulations refer unless
otherwise stated to formulations containing either the combination or a
component compound thereof. It will be understood that the administration
of the combination of the invention by means of a single patient pack, or
patient packs of each formulation, within a package insert diverting the
patient to the correct use of the invention is a desirable additional
feature of this invention. The invention also includes a double pack
comprising in association for separate administration, formulations of
tenofovir disoproxil fumarate and emtricitabine, or a physiologically
functional derivative of either or both thereof.

[0068] The combination therapies of the invention include: (1) a
combination of tenofovir DF and emtricitabine or (2) a combination
containing a physiologically functional derivative of either or both
thereof.

[0069] The combination may be formulated in a unit dosage formulation
comprising a fixed amount of each active pharmaceutical ingredient for a
periodic, e.g. daily, dose or subdose of the active ingredients.

[0070] Pharmaceutical formulations according to the present invention
comprise a combination according to the invention together with one or
more pharmaceutically acceptable carriers or excipients and optionally
other therapeutic agents. Pharmaceutical formulations containing the
active ingredient may be in any form suitable for the intended method of
administration. When used for oral use for example, tablets, troches,
lozenges, aqueous or oil suspensions, dispersible powders or granules,
emulsions, hard or soft capsules, syrups or elixirs may be prepared
(Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.).
Compositions intended for oral use may be prepared according to any
method known to the art for the manufacture of pharmaceutical
compositions and such compositions may contain one or more agents
including antioxidants, sweetening agents, flavoring agents, coloring
agents and preserving agents, in order to provide a palatable
preparation. Tablets containing the active ingredient in admixture with
non-toxic pharmaceutically acceptable excipient which are suitable for
manufacture of tablets are acceptable. These excipients may be, for
example, inert diluents, such as calcium or sodium carbonate, lactose,
lactose monohydrate, croscarmellose sodium, povidone, calcium or sodium
phosphate; granulating and disintegrating agents, such as maize starch,
or alginic acid; binding agents, such as cellulose, microcrystalline
cellulose, starch, gelatin or acacia; and lubricating agents, such as
magnesium stearate, stearic acid or talc. Tablets may be uncoated or may
be coated by known techniques including microencapsulation to delay
disintegration and absorption in the gastrointestinal tract and thereby
provide a sustained action over a longer period. For example, a time
delay material such as glyceryl monostearate or glyceryl distearate alone
or with a wax may be employed.

[0071] Formulations for oral use may be also presented as hard gelatin
capsules where the active ingredient is mixed with an inert solid
diluent, for example pregelatinized starch, calcium phosphate or kaolin,
or as soft gelatin capsules wherein the active ingredient is mixed with
water or an oil medium, such as peanut oil, liquid paraffin or olive oil.

[0072] Aqueous suspensions of the invention contain the active materials
in admixture with excipients suitable for the manufacture of aqueous
suspensions. Such excipients include a suspending agent, such as sodium
carboxymethylcellulose, methylcellulose, hydroxypropyl methylcelluose,
sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia, and
dispersing or wetting agents such as a naturally occurring phosphatide
(e.g., lecithin), a condensation product of an alkylene oxide with a
fatty acid (e.g., polyoxyethylene stearate), a condensation product of
ethylene oxide with a long chain aliphatic alcohol (e.g.,
heptadecaethyleneoxycetanol), a condensation product of ethylene oxide
with a partial ester derived from a fatty acid and a hexitol anhydride
(e.g., polyoxyethylene sorbitan monooleate). The aqueous suspension may
also contain one or more preservatives such as ethyl or n-propyl
p-hydroxybenzoate, one or more coloring agents, one or more flavoring
agents and one or more sweetening agents, such as sucrose, sucralose or
saccharin.

[0073] Oil suspensions may be formulated by suspending the active
ingredient in a vegetable oil, such as arachis oil, olive oil, sesame oil
or coconut oil, or in a mineral oil such as liquid paraffin. The oral
suspensions may contain a thickening agent, such as beeswax, hard
paraffin or cetyl alcohol. Sweetening agents, such as those set forth
above, and flavoring agents may be added to provide a palatable oral
preparation. These compositions may be preserved by the addition of an
antioxidant such as ascorbic acid, BHT, etc.

[0074] Dispersible powders and granules of the invention suitable for
preparation of an aqueous suspension by the addition of water provide the
active ingredient in admixture with a dispersing or wetting agent, a
suspending agent, and one or more preservatives. Suitable dispersing or
wetting agents and suspending agents are exemplified by those disclosed
above. Additional excipients, for example sweetening, flavoring and
coloring agents, may also be present.

[0075] The pharmaceutical compositions of the invention may also be in the
form of oil-in-water emulsions or liposome formulations. The oily phase
may be a vegetable oil, such as olive oil or arachis oil, a mineral oil,
such as liquid paraffin, or a mixture of these. Suitable emulsifying
agents include naturally-occurring gums, such as gum acacia and gum
tragacanth, naturally occurring phosphatides, such as soybean lecithin,
esters or partial esters derived from fatty acids and hexitol anhydrides,
such as sorbitan monooleate, and condensation products of these partial
esters with ethylene oxide, such as polyoxyethylene sorbitan monooleate.
The emulsion may also contain sweetening and flavoring agents. Syrups and
elixirs may be formulated with sweetening agents, such as glycerol,
sorbitol or sucrose. Such formulations may also contain a demulcent, a
preservative, a flavoring or a coloring agent.

[0076] The pharmaceutical compositions of the invention may be in the form
of a sterile injectable preparation, such as a sterile injectable aqueous
or oleaginous suspension. This suspension may be formulated according to
the known art using those suitable dispersing or wetting agents and
suspending agents which have been mentioned above. The sterile injectable
preparation may also be a sterile injectable solution or suspension in a
non-toxic parenterally acceptable diluent or solvent, such as a solution
in 1,3-butane-diol or prepared as a lyophilized powder. Among the
acceptable vehicles and solvents that may be employed are water, Ringer's
solution and isotonic sodium chloride solution. In addition, sterile
fixed oils may conventionally be employed as a solvent or suspending
medium. For this purpose any bland fixed oil may be employed including
synthetic mono- or diglycerides. In addition, fatty acids such as oleic
acid may likewise be used in the preparation of injectables.

[0077] The pharmaceutical compositions of the invention may be injected
parenterally, for example, intravenously, intraperitoneally,
intrathecally, intraventricularly, intrastemally, intracranially,
intramuscularly or subcutaneously, or they may be administered by
infusion techniques. They are best used in the form of a sterile aqueous
solution which may contain other substances, for example, enough salts or
glucose to make the solution isotonic with blood. The aqueous solutions
should be suitably buffered (preferably to a pH of from 3 to 9), if
necessary. The preparation of suitable parenteral formulations under
sterile conditions is readily accomplished by standard pharmaceutical
techniques well known to those skilled in the art.

[0078] The pharmaceutical compositions of the invention may also be
administered intranasally or by inhalation and are conveniently delivered
in the form of a dry powder inhaler or an aerosol spray presentation from
a pressurized container or a nebuliser with the use of a suitable
propellant, e.g. dichlorodifluoromethane, trichlorofluoromethane,
dichlorotetrafluoroethane, a hydrofluoroalkane such as
1,1,1,2-tetrafluoroethane (HFC 134a), carbon dioxide or other suitable
gas. In the case of a pressurized aerosol, the dosage unit may be
determined by providing a valve to deliver a metered amount. The
pressurized container or nebuliser may contain a solution or suspension
of the composition, e.g. using a mixture of ethanol and the propellant as
the solvent, which may additional contain a lubricant, e.g. sorbitan
trioleate. Capsules and cartridges (made, for example, from gelatin) for
use in an inhaler or insufflator may be formulated to contain a powder
mix of a compound of the formula (I) and a suitable powder base such as
lactose or starch. Aerosol or dry powder formulations are preferably
arranged so that each metered dose or "puff" contains from 20 μg to 20
mg of a composition for delivery to the patient. The overall daily dose
with an aerosol will be in the range of from 20 μg to 20 mg which may
be administered in a single dose or, more usually, in divided doses
throughout the day.

[0079] The amount of active ingredient that may be combined with the
carrier material to produce a single dosage form will vary depending upon
the host treated and the particular mode of administration. For example,
a time-release formulation intended for oral administration to humans may
contain approximately 1 to 1000 mg of active material compounded with an
appropriate and convenient amount of carrier material which may vary from
about 5 to about 95% of the total compositions (weight:weight). The
pharmaceutical composition can be prepared to provide easily measurable
amounts for administration. For example, an aqueous solution intended for
intravenous infusion may contain from about 3 to 500 μg of the active
ingredient per milliliter of solution in order that infusion of a
suitable volume at a rate of about 30 mL/hr can occur. As noted above,
formulations of the present invention suitable for oral administration
may be presented as discrete units such as capsules, cachets or tablets
each containing a predetermined amount of the active ingredient; as a
powder or granules; as a solution or a suspension in an aqueous or
non-aqueous liquid; or as an oil-in-water liquid emulsion or a
water-in-oil liquid emulsion. The active ingredient may also be
administered as a bolus, electuary or paste.

[0080] The combinations of the invention may conveniently be presented as
a pharmaceutical formulation in a unitary dosage form. A convenient
unitary dosage formulation contains the active ingredients in any amount
from 1 mg to 1 g each, for example but not limited to, 10 mg to 300 mg.
The synergistic effects of tenofovir DF in combination with emtricitabine
may be realized over a wide ratio, for example 1:50 to 50:1 (tenofovir
DF:emtricitabine). In one embodiment, the ratio may range from about 1:10
to 10:1. In another embodiment, the weight/weight ratio of tenofovir to
emtricitabine in a co-formulated combination dosage form, such as a pill,
tablet, caplet or capsule will be about 1, i.e. an approximately equal
amount of tenofovir DF and emtricitabine. In other exemplary
co-formulations, there may be more or less tenofovir than FTC. For
example, 300 mg tenofovir DF and 200 mg emtricitabine can be
co-formulated in a ratio of 1.5:1 (tenofovir DF: emtricitabine). In one
embodiment, each compound will be employed in the combination in an
amount at which it exhibits antiviral activity when used alone. Exemplary
Formulations A, B, C, D, E, and F (Examples) have ratios of 12:1 to 1:1
(tenofovir DF emtricitabine). Exemplary Formulations A, B, C, D, E, and F
use amounts of tenofovir DF and emtricitabine ranging from 25 mg to 300
mg. Other ratios and amounts of the compounds of said combinations are
contemplated within the scope of the invention.

[0081] A unitary dosage form may further comprise tenofovir DF and
emtricitabine, or physiologically functional derivatives of either
thereof, and a pharmaceutically acceptable carrier.

[0082] It will be appreciated by those skilled in the art that the amount
of active ingredients in the combinations of the invention required for
use in treatment will vary according to a variety of factors, including
the nature of the condition being treated and the age and condition of
the patient, and will ultimately be at the discretion of the attending
physician or health care practitioner. The factors to be considered
include the route of administration and nature of the formulation, the
animal's body weight, age and general condition and the nature and
severity of the disease to be treated. For example, in a Phase I/II
monotherapy study of emtricitabine, patients received doses ranging from
25 mg to 200 mg twice-a-day for two weeks. At each dose regimen greater
or equal to 200 mg, a 98-percent (1.75 log 10) or greater viral
suppression was observed. A once-a-day dose of 200 mg of emtricitabine
reduced the viral load by an average of 99 percent (1.92 log 10).
Viread® (tenofovir DF) has been approved by the FDA for the treatment
and prophylaxis of HIV infection as a 300 mg oral tablet. Emtriva®
(emtricitabine) has been approved by the FDA for the treatment of HIV as
a 200 mg oral tablet.

[0083] It is also possible to combine any two of the active ingredients in
a unitary dosage form for simultaneous or sequential administration with
a third active ingredient. The three-part combination may be administered
simultaneously or sequentially. When administered sequentially, the
combination may be administered in two or three administrations. Third
active ingredients have anti-HIV activity and include protease inhibitors
(PI), nucleoside reverse transcriptase inhibitors (NRTI), non-nucleoside
reverse transcriptase inhibitors (NNRTI), and integrase inhibitors.
Exemplary third active ingredients to be administered in combination with
tenofovir DF, emtricitabine, and their physiological functional
derivatives, are shown in Table A.

[0084] Another aspect of the present invention is a three-part combination
comprising tenofovir DF, FTC, and
9-[(R)-2-[[(S)-[[(S)-1-(isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]-
methoxy]propyl]adenine, also designated herein as GS-7340, which has the
structure:

[0086] For example, a ternary unitary dosage may contain 1 mg to 1000 mg
of tenofovir disoproxil fumarate, 1 mg to 1000 mg of emtricitabine, and 1
mg to 1000 mg of the third active ingredient. As a further feature of the
present invention, a unitary dosage form may further comprise tenofovir
DF, emtricitabine, the third active ingredient, or physiologically
functional derivatives of the three active ingredients thereof, and a
pharmaceutically acceptable carrier.

[0087] Combinations of the present invention enable patients greater
freedom from multiple dosage medication regimens and ease the needed
diligence required in remembering and complying with complex daily dosing
times and schedules. By combining tenofovir disoproxil fumarate and
emtricitabine into a single dosage form, the desired daily regimen may be
presented in a single dose or as two or more sub-doses per day. The
combination of co-formulated tenofovir DF and emtricitabine may be
administered as a single pill, once per day.

[0088] A further aspect of the invention is a patient pack comprising at
least one active ingredient: tenofovir disoproxil fumarate,
emtricitabine, or a physiologically functional derivative of either of
the combination and an information package or product insert containing
directions on the use of the combination of the invention.

[0089] Segregation of active ingredients in pharmaceutical powders and
granulations is a widely recognized problem that can result in
inconsistent dispersions of the active ingredients in final dosage forms.
Some of the main factors contributing to segregation are particle size,
shape and density. Segregation is particularly troublesome when
attempting to formulate a single homogenous tablet containing multiple
active ingredients having different densities and different particle
sizes. Glidants are substances that have traditionally been used to
improve the flow characteristics of granulations and powders by reducing
interparticulate friction. See Lieberman, Lachman, & Schwartz,
Pharmaceutical Dosage Forms: Tablets, Volume 1, p. 177-178 (1989),
incorporated herein by reference. Glidants are typically added to
pharmaceutical compositions immediately prior to tablet compression to
facilitate the flow of granular material into the die cavities of tablet
presses. Glidants include: colloidal silicon dioxide, asbestos free talc,
sodium aluminosilicate, calcium silicate, powdered cellulose,
microcrystalline cellulose, corn starch, sodium benzoate, calcium
carbonate, magnesium carbonate, metallic stearates, calcium stearate,
magnesium stearate, zinc stearate, stearowet C, starch, starch 1500,
magnesium lauryl sulfate, and magnesium oxide. Exemplary Tablet
Formulation A has colloidal silicon dioxide (Examples). Glidants can be
used to increase and aid blend composition homogeneity in formulations of
anti-HIV drugs (U.S. Pat. No. 6,113,920). The novel compositions of the
present invention may contain glidants to effect and maintain homogeneity
of active ingredients during handling prior to tablet compression.

[0090] The present invention provides pharmaceutical formulations
combining the active ingredients tenofovir DF and emtricitabine, or
physiologically functional derivatives thereof, in a sufficiently
homogenized form, and a method for using this pharmaceutical formulation.
An object of the present invention is to utilize glidants to reduce the
segregation of active ingredients in pharmaceutical compositions during
pre-compression material handling. Another object of the present
invention is to provide a pharmaceutical formulation combining the active
ingredients tenofovir DF and emtricitabine, or physiologically functional
derivatives thereof, with a pharmaceutically acceptable glidant,
resulting in a mixture characterized by a pharmaceutically acceptable
measure of homogeneity.

[0091] Formulations include those suitable for oral, rectal, nasal,
topical (including transdermal, buccal and sublingual), vaginal or
parenteral (including subcutaneous, intramuscular, intravenous and
intradermal) administration. The formulations may conveniently be
presented in unit dosage form and may be prepared by any methods well
known in the art of pharmacy. Such methods represent a further feature of
the present invention and include the step of bringing into association
the active ingredients with the carrier, which constitutes one or more
accessory ingredients, and maintaining chemical stability. In general,
the formulations are prepared by uniformly and intimately bringing into
association the active ingredients with liquid carriers or finely divided
solid carriers or both, and then if necessary shaping the product.

[0092] Formulations of the present invention suitable for oral
administration may be presented as discrete units such as capsules,
caplets, cachets or tablets each containing a predetermined amount of the
active ingredients; as a powder or granules; as a solution or a
suspension in an aqueous or non-aqueous liquid; or as an oil-in-water
liquid emulsion or a water-in-oil liquid emulsion. The active ingredient
may also be presented as a bolus, electuary or paste.

[0093] A tablet may be made by compression or molding, optionally with one
or more accessory ingredients. Compressed tablets may be prepared by
compressing in a suitable machine the active ingredients in a
free-flowing form such as a powder or granules, optionally mixed with a
binder (e.g. povidone, gelatin, hydroxypropyl methylcellulose),
lubricant, inert diluent, preservative, disintegrant (e.g. sodium starch
glycollate, cross-linked povidone, cross-linked sodium carboxymethyl
cellulose) surface-active or dispersing agent. Molded tablets may be made
by molding a mixture of the powdered compound moistened with an inert
liquid diluent in a suitable machine. The tablets may optionally be
coated or scored and may be formulated so as to provide slow or
controlled release of the active ingredients therein using, for example,
cellulose ether derivatives (e.g., hydroxypropyl methylcellulose) or
methacrylate derivatives in varying proportions to provide the desired
release profile. Tablets may optionally be provided with an enteric
coating, to provide release in parts of the gut other than the stomach.

[0094] Formulations suitable for topical administration in the mouth
include lozenges comprising the active ingredients in a flavored base,
usually sucrose and acacia or tragacanth; pastilles comprising the active
ingredient in an inert basis such as gelatin and glycerin, or sucrose and
acacia; and mouthwashes comprising the active ingredient in a suitable
liquid carrier. Formulations for rectal administration may be presented
as a suppository with a suitable base comprising, for example, cocoa
butter or a salicylates. Topical administration may also be by means of a
transdermal iontophoretic device.

[0095] Formulations suitable for vaginal administration may be presented
as pessaries, tampons, creams, gels, pastes, foams or spray formulations
containing in addition to the active ingredient such carriers as are
known in the art to be appropriate.

[0096] Formulations suitable for penile administration for prophylactic or
therapeutic use may be presented in condoms, creams, gels, pastes, foams
or spray formulations containing in addition to the active ingredient
such carriers as are known in the art to be appropriate.

[0097] Pharmaceutical formulations suitable for rectal administration
wherein the carrier is a solid are most preferably presented as unit dose
suppositories. Suitable carriers include cocoa butter and other materials
commonly used in the art. The suppositories may be conveniently formed by
admixture of the active combination with the softened or melted
carrier(s) followed by chilling and shaping in moulds.

[0098] Formulations suitable for parenteral administration include aqueous
and nonaqueous isotonic sterile injection solutions which may contain
anti-oxidants, buffers, bacteriostats and solutes which render the
formulation isotonic with the blood of the intended recipient; and
aqueous and non-aqueous sterile suspensions which may include suspending
agents and thickening agents; and liposomes or other microparticulate
systems which are designed to target the compound to blood components or
one or more organs. The formulations may be presented in unit-dose or
multi-dose sealed containers, for example, ampoules and vials, and may be
stored in a freeze-dried (lyophilized) condition requiring only the
addition of the sterile liquid carrier, for example water for injection,
immediately prior to use. Extemporaneous injection solutions and
suspensions may be prepared from sterile powders, granules and tablets of
the kind previously described.

[0099] Exemplary unit dosage formulations are those containing a daily
dose or daily subdose of the active ingredients, as hereinbefore recited,
or an appropriate fraction thereof. It should be understood that in
addition to the ingredients particularly mentioned above the formulations
of this invention may include other agents conventional in the art having
regard to the type of formulation in question, for example, those
suitable for oral administration may include such further agents as
sweeteners, thickeners and flavoring agents.

[0100] The compounds of the combination of the present invention may be
obtained in a conventional manner, known to those skilled in the art.
Tenofovir disoproxil fumarate can be prepared, for example, as described
in U.S. Pat. No. 5,977,089. Methods for the preparation of FTC are
described in WO 92/14743, incorporated herein by reference.

Composition Use

[0101] Compositions of the present invention are administered to a human
or other mammal in a safe and effective amount as described herein. These
safe and effective amounts will vary according to the type and size of
mammal being treated and the desired results of the treatment. Any of the
various methods known by persons skilled in the art for packaging
tablets, caplets, or other solid dosage forms suitable for oral
administration, that will not degrade the components of the present
invention, are suitable for use in packaging. The combinations may be
packaged in glass and plastic bottles. Tablets, caplets, or other solid
dosage forms suitable for oral administration may be packaged and
contained in various packaging materials optionally including a
dessicant, e.g. silica gel. Packaging may be in the form of unit dose
blister packaging. For example, a package may contain one blister tray of
tenofovir DF and another blister tray of emtricitabine pills, tablets,
caplets, or capsule. A patient would take one dose, e.g. a pill, from one
tray and one from the other. Alternatively, the package may contain a
blister tray of the co-formulated combination of tenofovir DF and
emtricitabine in a single pill, tablet, caplet or capsule. As in other
combinations and packaging thereof, the combinations of the invention
include physiological functional derivatives of tenofovir DF and FTC.

[0102] The packaging material may also have labeling and information
related to the pharmaceutical composition printed thereon. Additionally,
an article of manufacture may contain a brochure, report, notice,
pamphlet, or leaflet containing product information. This form of
pharmaceutical information is referred to in the pharmaceutical industry
as a "package insert." A package insert may be attached to or included
with a pharmaceutical article of manufacture. The package insert and any
article of manufacture labeling provides information relating to the
pharmaceutical composition. The information and labeling provides various
forms of information utilized by health-care professionals and patients,
describing the composition, its dosage and various other parameters
required by regulatory agencies such as the United States Food and Drug
Agency.

Assays of the Combinations

[0103] The combinations of the inventions may be tested for in vitro
activity against HIV and sensitivity, and for cytotoxicity in laboratory
adapted cell lines, e.g. MT2 and in peripheral blood mononuclear cells
(PBMC) according to standard assays developed for testing anti-HIV
compounds, such as WO 02/068058 and U.S. Pat. No. 6,475,491. Combination
assays may be performed at varying concentrations of the compounds of the
combinations to determine EC50 by serial dilutions.

Exemplary Formulations

[0104] The following examples further describe and demonstrate particular
embodiments within the scope of the present invention. Techniques and
formulations generally are found in Remington's Pharmaceutical Sciences
(Mack Publishing Co., Easton, Pa.). The examples are given solely for
illustration and are not to be construed as limitations as many
variations are possible without departing from spirit and scope of the
Invention. The following examples are intended for illustration only and
are not intended to limit the scope of the invention in any way. "Active
ingredient" denotes tenofovir disoproxil fumarate, emtricitabine, or a
physiologically functional derivative of either thereof.

Tablet Formulation

[0105] The following exemplary formulations A, B, C, D, E, and F are
prepared by wet granulation of the ingredients with an aqueous solution,
addition of extragranular components and then followed by addition of
magnesium stearate and compression.

[0114] The following controlled release capsule formulation is prepared by
extruding ingredients a, b, and c using an extruder, followed by
spheronization of the extrudate and drying. The dried pellets are then
coated with release-controlling membrane (d) and filled into a two-piece,
hard gelatin or hydroxypropyl methylcellulose capsule.

[0116] One-fifth of the Witepsol H15 is melted in a steam jacketed pan at
45° C. maximum. The active ingredients are sifted through a 200
micron sieve and added to the molten base with mixing, using a Silverson
fitted with a cutting head, until a smooth dispersion is achieved.
Maintaining the mixture at 45° C., the remaining Witepsol H15 is
added to the suspension and stirred to ensure a homogenous mix. The
entire suspension is passed through a 250 micron stainless steel screen
and, with continuous stirring, is allowed to cool to 40° C. At a
temperature of 38° C. to 40° C., 2.02 g of the mixture is
filled into suitable, 2 ml plastic molds. The suppositories are allowed
to cool to room temperature.

[0118] The effects of granulation water level (ranging from 40% to 50%
w/w) and wet massing time were studied on the physicochemical properties
of the final powder blend and its performance with respect to blend
uniformity and compressibility (tablet compactibility). In addition,
content uniformity, assay, stability and dissolution performance was
evaluated for the TDF/emtricitabine fixed dose combination tablets.

Formulation Equipment

[0119] Equipment included a high shear mixer equipped with a pressure tank
and spray nozzle tip to add the granulating water, a fluid-bed dryer, a
mill, a tumble blender, a rotary tablet press, and a tablet deduster.

Formulation Process

[0120] The dried, milled powder was blended with the extragranular
microcrystalline cellulose and croscarmellose sodium and then blended
with magnesium stearate. Powder samples were removed after mixing with
the magnesium stearate. The blend samples were evaluated for, bulk
density, mesh analysis and compressibility. The powder blend mixed with
the magnesium stearate was compressed into tablets on a press setup.

Materials

[0121] The following Table 1 lists the quantitative composition of the
TDF/emtricitabine tablet formulation.

[0122] Moisture content was measured by loss on drying using a heat
lamp/balance system. The powder blend was sampled with a sampling thief
fitted with chambers to determine powder blend uniformity. Duplicate
samples were removed from each of several locations in the blender. Blend
uniformity analysis was performed on one sample from each location.

[0123] Particle size analysis of the final powder blend was determined by
sifting a multi-gram sample through a screen using a sonic sifter. The
quantity of final powder blend retained on each sieve and the fines
collector was determined by calculating the difference in weight between
the sieves and fines collector before and after the test. The geometric
mean diameter particle size was calculated by logarithmic weighting of
the sieved distribution.

[0124] Bulk density was determined by filling a graduated cylinder with
the final powder blend and measuring the weight differential between the
empty and filled graduate cylinder per unit volume.

[0125] Tablets were characterized for friability using a friabilator, a
hardness tester, a thickness micrometer equipped with a printer, and a
weighing balance.

[0126] Compression characteristics were determined using a rotary tablet
press equipped with a flat-faced, beveled edged punch to a target weight
of 400 mg. The powder blends were compressed using target upper punch
pressures ranging from approximately 100 to 250 MPa. The apparent
normalized ejection force was determined and normalized for tablet
thickness and diameter.

[0127] Tablet hardness was determined using a hardness tester. Tablet
thickness was determined using a micrometer, and tablet weights were
determined using a top loading balance.

Wet Granulation

[0128] The powders were blended in a granulator and then granulated using
water. The impeller and chopper speeds were kept constant in the blender
at a low setting during the granulation and wet massing operations. After
water addition, the impeller and chopper were stopped and the granulator
bowl was opened to observe the granulation consistency and texture. The
lid was closed and the wet massing phase was performed. Acceptable
granules had 40% w/w and 60% w/w water, respectively.

Wet Milling

[0129] To facilitate a uniform drying process, each wet granulation was
deagglomerated with a mill fitted with a screen and an impeller. The
milled wet granules were charged into a fluid-bed dryer immediately
following wet milling.

Fluid-Bed Drying

[0130] Milled wet granules were dried using an inlet air setpoint
temperature of about 70° C. and airflow of approximately 100 cfm.
The target LOD was about 1.0% with a range of not more than (NMT) 1.5%.
The total fluid-bed drying time ranged from 53 to 75 minutes. Final LOD
ranged from 0.4% to 0.7% for all of the batches dried. The final exhaust
temperatures for all the batches ranged from 47° C. to 50°
C.

Dry Milling

[0131] All dried granules were milled through a perforated screen. The
mill was equipped with a square impeller and operated. The lots were
milled and manually transferred to the V-blender.

Blending

[0132] Each lot was blended using the V-blender. In one set of three
formulations, starting with 12 kg materials, final powder blend yield
available for compression after blending ranged from 10.5 kg (87.5%) to
11.1 kg (92.5%). The final powder blend bulk density ranged from 0.48 to
0.58 g/cc and the geometric mean diameter particle size ranged from 112
to 221 μm. Percent water and wet massing time affect final powder
blend particle size and bulk density.

[0133] The powder blending for both tenofovir DF and emtricitabine gave a
mean (n=10) strength value for tenofovir DF ranged from 100.6% to 102.8%
of target strength for the lots and the relative standard deviation (RSD)
was from 0.5% to 1.7%. The mean (n=10) strength value for emtricitabine
ranged from 101.3% to 104.1% of target strength for the lots with the
relative standard deviation (RSD) ranged from 0.6% to 1.7%. The final
powder blend moisture level ranged from 0.8% to 1.1% LOD.

Tablet Compression

[0134] The final blends were compressed using a rotary tablet press and
the tablets were film-coated.

[0135] Three 300 gm formulations (Table 2) were granulated in a granulator
equipped with a 1-L bowl. The quantities of intragranular components were
based on a 300 g total batch size. The formulations in lots 1 and 2
differed in the amount of microcrystalline cellulose 30% vs. 20% w/w,
respectively. Lots 2 and 3 were identical except for the type of binder.
Lot 2 contained 5% w/w of pregelatinized starch and lot 3 contained 5%
w/w povidone as binder.

[0136] After water addition, the impeller and chopper were stopped and the
granulator bowl was opened to observe the granulation consistency and
texture. To achieve similar granulation consistency, lots 1, 2, and 3
were granulated with 45%, 40%, and 30% w/w water, respectively. The lid
was closed and the wet massing phase was performed. All lots had a 30 sec
wet massing resulting in acceptable granulations. The wet granulations
from all batches were hand screened through a sieve to deagglomerate. The
resulting granulations were tray dried in a convection oven set at
60° C. for approximately 20 hours to an LOD <1.0%. The dried
granulations from all batches were hand screened through a sieve. In
order to fit the granulation into the small scale (300 mL) V-blender, the
final blend batch size was adjusted to 100 g. A portion, 81 g of the
resulting blend from Lot 1 was blended with 15 g microcrystalline
cellulose, 3 g croscarmellose sodium and 1 g magnesium stearate. 86 g of
the resulting granulation from Lot 2 and Lot 3 were each blended with 10
g microcrystalline cellulose, 3 g croscarmellose sodium and 1 g magnesium
stearate.

[0137] Purity analysis was conducted by reverse-phase HPLC (high
performance liquid chromatography). Impurities related to tenofovir
disoproxil fumarate and emtricitabine were characterized and measured in
the bulk API (active pharmaceutical ingredient) before formulation in the
three lots of Table 2, and again after formulation in the resulting
tablets. The impurities include by-products from hydrolysis of the
exocyclic amino groups of tenofovir disoproxil fumarate and
emtricitabine, and the hydrolysis of the disoproxil (POC) esters of
tenofovir disoproxil fumarate. In each lot, the sum total of impurities
related to tenofovir disoproxil fumarate and emtricitabine was less than
1% after formulation and tablet manufacture.

[0138] The physicochemical properties of tenofovir disoproxil fumarate and
emtricitabine tablets were evaluated by visual appearance, water content,
label strength, impurity and degradation product contents, and tablet
dissolution. Stability studies were conducted on drug product packaged in
container-closure systems that are identical to the intended clinical and
commercial container-closure system. There was no sign of discoloration
or tablet cracking during the course of the stability study. Film-coated
tenofovir disoproxil fumarate and emtricitabine tablets exhibited
satisfactory stability at 40° C./75% RH (relative humidity) for up
to six months when packaged and stored with silica gel desiccant. No
significant loss (defined as ≧5% degradation) in % label strength
of tenofovir DF or emtricitabine was observed after six months at
40° C./75% RH. when packaged and stored with desiccant. The
increase in the total degradation products was 1.5% for tenofovir DF and
0.6-0.7% for emtricitabine after six months at 40° C./75% RH when
packaged and stored with 3 grams of desiccant.

[0139] All publications and patent applications cited herein are
incorporated by reference to the same extent as if each individual
publication or patent application was specifically and individually
indicated to be incorporated by reference.

[0140] Although certain embodiments are described in detail above, those
having ordinary skill in the art will clearly understand that many
modifications are possible in the claims without departing from the
teachings thereof. All such modifications are intended to be encompassed
within the claims of the invention.

EMBODIMENTS OF THE INVENTION

[0141] A1. A pharmaceutical composition comprising an effective amount of
a compound of the formula: